Fermentation processes are hostile environments for microorganisms such as yeast, subjecting them to numerous stresses. These stresses can lead to defective or sub-optimal fermentation in which yeast metabolism is halted and/or undesirable flavours are produced (Ivorra et. al., 1999).
While the major requirements for successful fermentation have been identified though previous research efforts, there are no recognised tools that allow the rapid detection of the source of fermentation problems early enough to circumvent them. Indeed, the majority of industrial fermentations are monitored purely by the assessment of the specific gravity of the fermentation medium. Accordingly, in an attempt to overcome or at least alleviate some of the problems associated with monitoring fermentation the applicant proposes utilising molecular techniques to identify genes which are indicative of sub-optimal fermentation. In particular the applicant proposes using molecular methods to monitor the in vivo effects of varying fermentation conditions so that a more accurate assessment of the effect of these conditions on the microorganisms present during fermentation can be undertaken.
Until recently the rate limiting step in the development of methods for monitoring fermentation was the identification of suitable genes. However, the applicant has now found that by using genome-wide transcriptional analysis, genes can be identified that meet the methods' requirements. Therefore, the applicant proposes that the use of this technique can readily determine which genes are highly responsive and specific to defined stresses, such as zinc-limiting conditions. Having identified potentially useful genes, their “normal” levels of expression can be determined so that an accurate assessment of sub-optimal expression levels can be identified during fermentation.
Genes that may be of use include HSP12 (Ivorra et. al., 1999) and SPI1 (Puig and Perez-Ortin, 2000) which have been shown to have promise as genetic markers for stress conditions in alcoholic fermentation. While these genes respond to a broad range of sub-optimal conditions and allow the identification of the presence of stress per se, their non-specificity prevents their use to identify the cause or type of stress.
Zinc deficiency, for instance, is a major contributor to the retardation of yeast fermentation in the beer brewing process (Bromberg et. al., 1997). The lack of available zinc slows down the fermentation rate, even leading to the complete cessation of fermentation or a “stuck brew”. The direct measurement of zinc levels in wort is not a reliable measure of zinc deficiency, since zinc can be present in a form that is not available to the yeast.
The applicant has accordingly further identified a gene whose expression levels are directly affected by the presence of zinc. The applicant proposes that this gene will be useful for monitoring a fermentation process for zinc deficiency.
It will be appreciated by those skilled in the art that there are a number of techniques that can be used to determine the level of gene expression. For example, Northern hybridisation analysis, dot blots and real-time polymerase chain reaction.
It will be clearly understood that, although a number of prior art publications are referred to herein, this reference does not constitute an admission that any of these documents form part of the common general knowledge in the art, in Australia or in any other country.